N-RAP in Dilated Cardiomyopathy: The muscle-specific protein N-RAP is concentrated at cardiac intercalated disks, plays a role in myofibril assembly, and is upregulated early in mouse models of dilated cardiomyopathy. Using a tet-off system, we developed novel transgenic lines exhibiting cardiac-specific N-RAP overexpression 2.5 times greater than normal. At 40-50 weeks, N-RAP overexpression resulted in dilation and decreased ejection fraction in both ventricles. Expression of transcripts encoding brain natriuretic peptide and skeletal alpha-actin was increased by cardiac-specific N-RAP overexpression, indicative of a cardiomyopathic response. N-RAP overexpression did not alter the levels or organization of N-cadherin and connexin-43. The results show that chronic N-RAP overexpression in the mouse leads to dilated cardiomyopathy by 10 months, and that the early N-RAP upregulation previously observed in some mouse models of dilated cardiomyopathy is unlikely to account for the remodeling of intercalated disks observed in those cases. Instead, N-RAP upregulation and intercalated disk remodeling are likely to be parts of two distinct pathways leading to cardiomyopathy, and their combined effects may cause a more severe dilated cardiomyopathy phenotype than either alone. Myofibril Function in Pompe Disease: Pompe disease, a deficiency of lysosomal acid alpha-glucosidase, is a disorder of glycogen metabolism that can affect infants, children, or adults. In all forms of the disease, there is progressive muscle pathology leading to premature death. The pathology is characterized by accumulation of glycogen in lysosomes, autophagic buildup, and muscle atrophy. The purpose of the present investigation was to determine if myofibrillar dysfunction in Pompe disease contributes to muscle weakness beyond that attributed to atrophy. The study was performed on isolated myofibers dissected from severely affected fast glycolytic muscle in the alpha-glucosidase knockout mouse model. Psoas muscle fibers were first permeabilized, so that the contractile proteins could be directly relaxed or activated by control of the composition of the bathing solution. When normalized by cross-sectional area, single fibers from knockout mice produced 6.3 N/cm2 of maximum calcium-activated tension compared with 12.0 N/cm2 produced by wild-type fibers. The total protein concentration was slightly higher in the knockout mice, but concentrations of the contractile proteins myosin and actin remained unchanged. Structurally, X-ray diffraction showed that the actin and myosin filaments, normally arranged in hexagonal arrays, were disordered in the knockout muscle, and a lower fraction of myosin cross bridges was near the actin filaments in the relaxed muscle. The results are consistent with a disruption of actin and myosin interactions in the knockout muscles, demonstrating that impaired myofibrillar function contributes to weakness in the diseased muscle fibers.